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Published on September 18, 2003; 10.1104/pp.103.024554

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Received March 31, 2003
Returned for revision May 8, 2003
Accepted July 1, 2003

Down-Regulating {alpha}-Galactosidase Enhances Freezing Tolerance in Transgenic Petunia

Joyce C. Pennycooke , Michelle L. Jones , and Cecil Stushnoff *

Department of Horticulture and Landscape Architecture, Colorado State University, Fort Collins, Colorado 80523 (J.C.P., C.S.); and Department of Horticulture and Crop Science, The Ohio State University/Ohio Research and Development Center, Wooster, Ohio 44691 (M.L.J.)

* Corresponding author; email: stushnof{at}lamar.colostate.edu.

{alpha}-Galactosidase ({alpha}-Gal; EC 3.2.1.22) is involved in many aspects of plant metabolism, including hydrolysis of the {alpha}-1,6 linkage of raffinose oligosaccharides during deacclimation. To examine the relationship between endogenous sugars and freezing stress, the expression of {alpha}-Gal was modified in transgenic petunia (Petunia x hybrida cv Mitchell). The tomato (Lycopersicon esculentum) Lea-Gal gene under the control of the Figwort Mosaic Virus promoter was introduced into petunia in the sense and antisense orientations using Agrobacterium tumefaciens-mediated transformation. RNA gel blots confirmed that {alpha}-Gal transcripts were reduced in antisense lines compared with wild type, whereas sense plants had increased accumulation of {alpha}-Gal mRNAs. {alpha}-Gal activity followed a similar trend, with reduced activity in antisense lines and increased activity in all sense lines evaluated. Raffinose content of nonacclimated antisense plants increased 12- to 22-fold compared with wild type, and 22- to 53-fold after cold acclimation. Based upon electrolyte leakage tests, freezing tolerance of the antisense lines increased from -4°C for cold-acclimated wild-type plants to -8°C for the most tolerant antisense line. Down-regulating {alpha}-Gal in petunia results in an increase in freezing tolerance at the whole-plant level in nonacclimated and cold-acclimated plants, whereas overexpression of the {alpha}-Gal gene caused a decrease in endogenous raffinose and impaired freezing tolerance. These results suggest that engineering raffinose metabolism by transformation with {alpha}-Gal provides an additional method for improving the freezing tolerance of plants.




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